In 1895, Roentgen (Nature 53:274-276 (1896)) discovered X-rays and their use to visualize bones and organs in a living body. In a typical experiment/diagnosis, X-rays would be directed to the patient and the resulting image would be collected on a film residing behind the patient. The developed film would show a qualitative picture of where the X-rays passed through the body. For example, soft tissues appear darker than dense structures such as bones, which absorb more of the X-rays. It was not until the 1970's, however, with the advent of computer technology coupled with X-ray technology, that this became a breakthrough in medical imaging (i.e., X-ray Computer Tomography; CT) (G. N. Hounsfield British Pat. 1283915; Am. J. Radiol. 131:103 (1978)). By using mathematical methods/models developed by A. M. Cormack (J. Appl. Physics 34:2722-2727 (1978)) one can reconstruct an image of the tissues that the X-rays have passed through. This ability to map tissue density (i.e., X-ray attention) allowed X-ray CT to become a common and routine medical diagnostic technique used world-wide today.
Contrast agents for X-rays have been used for a number of years. One of the first and by far one of the most extensively used X-ray contrast agents are barium salts. Barium salts are typically used for gastrointestinal imaging. Besides barium salts other radiopaque compounds are known. Essentially any organic molecule with one or more iodines or bromines will attenuate X-rays. This inherent property of the bromines and iodines allows compounds containing such atoms to be used as CT contrast agent. One particular class of CT contrast agents are brominated fluorocarbons such as perfluorooctylbromide (PFOB).
Perfluorooctylbromide has been effectively used in a number of indications as a CT contrast agent including: 1) determination of acute renal and hepatic microvascular volumes in acute renal failure (Hillman et al., Invest Radiol. 17:41-45 (1982)); 2) a liver/spleen specific tumor imaging agent (Mattrey et al., Radiology 145:755-758 (1982); Patronas et al., Invest Radio. 19:570-573 (1984)); 3) PFOB blood pool contrast agent with imaging of the kidneys, liver, spleen, and mediastinum (Mattrey et al., J. Comput. Assist. Tomogr. 8:739-744 (1984); Peck et al., Invest. Radiol. 19:129-132 (1984)); 4) enhancement of liver abscesses with PFOB (Mattrey, R. F., Invest. Radio. 19:438-446, (1984); Mattrey et al., Invest. Radiol. 26:792-798 (1991); Adam et Invest. Radial. 27:698-705 (1992)); 5) hepatosplenic computed tomography in humans with PFOB (Bruneton et al., Invest. Radiol. 23:306-307 (1988)); 6) determination of liver metastatic cancer in humans (Bruneton et al., Radiology 170:179-183 (1989)); 7) bronchiolography with PFOB (Stern et al., J. Thorac. Imaging 8:300-304 (1993)); and 8) GI imaging with PFOB (Mattrey et al., Invest. Radiol. 26:65-71 (1991)).
Indeed, contrast agents are desirable in radiological imaging because they enhance visualization of the organs (i.e., their location, size and conformation) and other cellular structures from the surrounding medium. The soft tissues, for example, have similar cell composition (i.e., they are primarily composed of water) even though they may have remarkably different biological functions (e.g., liver and pancreas).
The technique of magnetic resonance imaging (MRI) or nuclear magnetic resonance (NMR) imaging relies on the detection of certain atomic nuclei at an applied magnetic field strength using radio-frequency radiation. In some respects it is similar to X-ray computer tomography (CT), in that it can provide (in some cases) cross-sectional images of organs with potentially excellent soft tissue resolution. In its current use, the images constitute a distribution map of protons in organs and tissues. However, unlike X-ray computer tomography, MRI does not use ionizing radiation. MRI is, therefore, a safe non-invasive technique for medical imaging.
While the phenomenon of NMR was discovered in 1954, it is only recently that it has found use in medical diagnostics as a means of mapping internal structure. The technique was first developed by Lauterbur (Nature 242:190-191 (1973)).
It is well known that nuclei with the appropriate nuclear spin align in the direction of the applied magnetic field. The nuclear spin may be aligned in either of two ways: with or against the external magnetic field. Alignment with the field is more stable; while energy must be absorbed to align in the less stable state (i.e. against the applied field). In the case of protons, these nuclei precess or resonate at a frequency of 42.6 MHz in the presence of a 1 tesla (1 tesla=104 gauss) magnetic field. At this frequency, a radio-frequency (RF) pulse of radiation will excite the nuclei and change their spin orientation to be aligned against the applied magnetic field. After the RF pulse, the excited nuclei "relax" or return to equilibrium or in alignment with the magnetic field. The decay of the relaxation signal can be described using two relaxation terms. T.sub.1, the spin-lattice relaxation time or longitudinal relaxation time, is the time required by the nuclei to return to equilibrium along the direction of the externally applied magnetic field. The second, T.sub.2, or spin-spin relaxation time, is associated with the dephasing of the initially coherent precession of individual proton spins. The relaxation times for various fluids, organs and tissues in different species of mammals is well documented.
One advantage of MRI is that different scanning planes and slice thicknesses can be selected without loss of resolution. This permits high quality transverse, coronal and sagittal images to be obtained directly. The absence of any mechanical moving parts in the MRI equipment promotes a high degree of reliability. It is generally believed that MRI has greater potential than X-ray computer tomography (CT) for the selective examination of tissues. In CT, the X-ray attenuation coefficients alone determine the image contrast, whereas at least three separate variables (T.sub.1, T.sub.2, and nuclear spin density) contribute to the magnetic resonance image.
Due to subtle physio-chemical differences among organs and tissue, MRI may be capable of differentiating tissue types and in detecting diseases that may not be detected by X-ray or CT. In comparison, CT and X-ray are only sensitive to differences in electron densities in tissues and organs. The images obtainable by MRI techniques can also enable a physician to detect structures smaller than those detectable by CT, due to its better spatial resolution. Additionally, any imaging scan plane can be readily obtained using MRI techniques, including transverse, coronal and sagittal.
Currently, MRI is widely used to aid in the diagnosis of many medical disorders. Examples include joint injuries, bone marrow disorders, soft tissue tumors, mediastinal invasion, lymphadenopathy, cavernous hemangioma, hemochromatosis, cirrhosis, renal cell carcinoma, uterine leiomyoma, adenomyosis, endometriosis, breast carcinomas, stenosis, coronary artery disease, aortic dissection, lipomatous hypertrophy, atrial septum, constrictive pericarditis, and the like (see, for example, Edelman & Warach, Medical Progress 328:708-716 (1993); Edelman & Warach, New England J. of Medicine 328:785-791 (1993)).
Routinely employed magnetic resonance images are presently based on proton signals arising from the water molecules within cells. Consequently, it is often difficult to decipher the images and distinguish individual organs and cellular structures. There are two potential means to better differentiate proton signals. The first involves using a contrast agent that alters the T.sub.1 or T.sub.2 of the water molecules in one region compared to another. For example, gadolinium diethylenetriaminepentaacetic acid (Gd-DTPA) shortens the proton T.sub.1 relaxation time of water molecules in near proximity thereto, thereby enhancing the obtained images.
Paramagnetic cations such as, for example, Gd, Mn, and Fe are excellent MRI contrast agents, as suggested above. Their ability to shorten the proton T.sub.1 relaxation time of the surrounding water enables enhanced MRI images to be obtained which otherwise would be unreadable.
The second route to differentiate the individual organs and cellular structures is to introduce another nucleus for imaging (i.e., an imaging agent). Using this second approach, imaging can only occur where the contrast agent has been delivered. An advantage of this method is the fact that imaging is achieved free from interference from the surrounding water. Suitable contrast agents must be bio-compatible (i.e. non-toxic, chemically stable, not reactive with tissues) and of limited lifetime before elimination from the body.
Although hydrogen has typically been selected as the basis for MRI scanning (because of its abundance in the body), this can result in poorly imaged areas due to lack of contrast. Thus the use of other active MRI nuclei (such as fluorine) can, therefore, be advantageous. The use of certain perfluorocarbons in various diagnostic imaging technologies such as ultrasound, magnetic resonance, radiography and computer tomography has been described in an article by Mattery (see SPIE, 626, XIV/PACS IV, 18-23 (1986)). The use of fluorine is advantageous since fluorine is not naturally found within the body.
Prior art suggestions of fluorine-containing compounds useful for magnetic resonance imaging for medical diagnostic purposes are limited to a select group of fluorine-containing molecules that are water soluble or can form emulsions. Accordingly, prior art use of fluorocarbon emulsions of aqueous soluble fluorocarbons suffers from numerous drawbacks, for example, 1) the use of unstable emulsions, 2) the lack of organ specificity and targeting, 3) the potential for inducing allergic reactions due to the use of emulsifiers and surfactants (e.g., egg phophatides and egg yolk lecithin), 4) limited delivery capabilities, and 5) water soluble fluorocarbons are quickly diluted in blood after intravenous injection.
Another medical imaging application for perfluorocarbon filled polymeric shells is ultrasonography. This non-invasive, non-iodizing radiation medical imaging technique is safe and currently used world-wide for a number of indications. Ultrasonic imaging (i.e., sonography) is based on the reflection of ultrasonic sound waves from an object. Thus, the acoustic properties of the material will have dramatic effects on the reflected (i.e., scattered) radiation or sound waves. The reflected or scattered ultrasound radiation is received by a probe that covers the area to be imaged. In a typical medical diagnosis, an ultrasonic transducer (ultrasonic frequency is typically in the MHz region) transmits ultrasound into a living body for which one wishes to obtain an image or diagnosis. The ultrasound travels through the region and scatters on structures (e.g., organs). This scattered ultrasound is then collected and an image is produced.
The magnitude of the reflected sound waves is dramatically dependent on the acoustic properties of the material. The acoustic properties of a substance depend both on the density as well as the velocity of the transmitted ultrasound. Materials typically have their greatest differences of acoustic properties at interfaces such as liquid-gas or liquid-solid. This difference in acoustic properties (i.e., the acoustic impedance) results in more intense reflected ultrasonic radiation. In a physical sense the reflected sound waves are influenced by the following: 1) the size of the scattering center, 2) the differences in density from the scattering center and the surrounding area, 3) the compressibility of the scattering center, and 4) the acoustic properties of the surrounding area. However, the scattered ultrasound that is received from an image and processed often lacks signal intensity, sharpness and clarity. Thus, contrast agents that will help to distinguish organs and tissues are of great need. By using materials that have different acoustic properties than the surrounding area it is possible to improve the resolution of the acquired image. One class of materials that have been used as ultrasonography contrast agents are perfluorohalocarbons.
Another medical imaging application for polymeric shells is electron paramagnetic resonance (EPR) imaging and spectroscopy. This non-invasive, non-iodizing radiation medical spectroscopy and imaging technique is safe and currently in preclinical development.
In order for EPR spectroscopy and imaging to be accomplished, the nitroxide free radical needs to be detected by the EPR instrument. However most nitroxide free radicals are not stable in vivo because they are bioreduced. This short half-life in vivo prevents this technique from being used for imaging. Yet, this technique has better sensitivity than magnetic resonance imaging and thus would be a medically useful technique. Accordingly, means to protect nitroxide free radicals from bioreduction by the in vivo environment would be of great value.